HighYield T7 RNA Crosslinking Kit (4-Thio-UTP), Synthesis of 4-Thio-modified RNA

Jena Bioscience

HighYield T7 RNA Crosslinking Kit (4-Thio-UTP), Synthesis of 4-Thio-modified RNA

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Research Use Only

HighYield T7 RNA Crosslinking Kit (4-Thio-UTP) is designed to produce randomly 4-Thio-modified RNA probes via in vitro transcription. Such probes are ideally suited for the detection of RNA-Protein interactions by photocrosslinking. The photoreactive Thio group triggers the formation of a covalent bond between the RNA and closely interacting proteins upon UV irradiation. 4-Thio-UTP is efficiently incorporated into RNA as substitute for its natural counterpart UTP using an optimized reaction buffer and T7 RNA Labeling Polymerase Mix. 80 % 4-Thio-UTP substitution typically results in an optimal balance between reaction and labeling efficiency. Individual optimization of 4-Thio-UTP/UTP ratio however, can easily be achieved with the single nucleotide format. The resulting 4-Thio-modified RNA probe can be used for photocrosslinking experiments using a 365 nm UV light. The kit contains sufficient reagents for 15 labeling reactions of 20 microl each (100 % 4-Thio-UTP substitution, 7.5 mM ATP, 7.5 mM GTP, 7.5 mM CTP,7.5 mM 4-Thio-UTP). A 20 microl reaction with 100 % substitution yields about 10 microg RNA, with 80 % substitution about 50 microg RNA after 30 min incubation (1 microg T7 control template, 1.4 kb RNA transcript). Yields may however vary depending on the template (promotor design, sequence length, secondary structure formation).

-20°C

41116133